Oral thrush (pseudomembranous candidiasis) continues to afflict an unacceptably high percentage of immunocompromised individuals, particularly children. Since the recalcitrant nature of this oral infection, triggered primarily by C. albicans, could be attributed to its biofilm forming activities, knowledge of the distinct stages in mucosal biofilm formation and concomitant host responses is of paramount importance in understanding the pathogenesis of this infection and designing effective anti-infective strategies. The goals of the proposed studies in the next competitive cycle are directed toward a better understanding of the host response to mucosal biofilms formed by Candida in the oral cavity and the role of proinflammatory cytokines and neutrophils in the prevention or elimination of such biofilms. In the current funding cycle we established a three-dimensional oral tissue model which faithfully reproduces the histologic and cellular characteristics of pseudomembranous candidiasis lesions and incorporated neutrophils so that we can examine their candidacidal activities in situ. While working with this tissue analogue we discovered that Candida does not interact with oral mucosal tissues as single cell organisms, but forms a complex mucosal biofilm. We were able to reproduce and validate the presence of a mucosal tissue biofilm in a mouse model of oroesophageal candidiasis. This application will build on data generated during the current funding cycle to address the following Aims: a) characterize the stages in development, cellular composition and spatial arrangement of biofilms in the oral mucosa and test the hypothesis that Candida tissue biofilms affect the mucosal proinflammatory cytokine response to infection;b) test the hypothesis that neutrophil anti-Candida responses are inhibited within the biofilm environment and examine the mechanisms of this inhibition;and c) investigate the role of specific Candida gene products in facilitating oral mucosal biofilm formation, tissue invasion and damage. Since superficial mycoses characterized by Candida biofilm formation on the surface of stratified epithelia affect other GI tract mucosal sites, such as the esophageal mucosa, which may be more vulnerable to invasion our studies may have far reaching implications in preventing disseminated infection. We envision that our studies will lead to the development of new oral anti-mycotic agents which target pathways of mucosal biofilm regulation by C. albicans and promote local neutrophil antifungal functions. Public Health Significance: Oral pseudomembranous candidiasis is still the most prevalent form of Candida infection in patients with weakened or immature immune systems, such as HIV+ children, neonates and patients with malignancies. We propose that in this infection biofilms provide a protective environment for Candida from innate defense mechanisms, which may promote its persistence. We envision that our studies in tissue biofilms will lead to the development of new oral anti-mycotic agents which target pathways of biofilm regulation by C. albicans and promote neutrophil antifungal functions.